This invention relates to a method of purifying protein and more specifically, to a more expedient and direct method for purifying C1-inhibitor, inhibitor of the first component of human complement (hereafter called C1-Inh).
C1-Inh controls C1 activation by forming covalent complexes with activated C1r and C1s in a one-to-one stoichiometry (Harpel and Cooper 1795, Reboul et al, 1977). C1-Inh can also form complexes with other target enzymes and thereby is involved in the regulation of several other plasma proteolytic systems including the coagulation, fibrinolytic and contact systems (Cooper 1985). The gravity of the disease associated with C1-Inh deficiency (hereditary angioedema) demonstrates the physiological importance of this plasma protein (Davis 1988).
Recently a new type of acquired C1-Inh deficiency characterized by the presence of autoantibodies to the C1-Inh molecule and an abnormally low molecular weight form of the C1-Inh has been described by several groups (Jackson et al, 1986; Alsenz et al, 1987; Malbran et al, 1988). The autoantibodies do not seem to interfere with the initial cleavage of the inhibitor by the target enzymes but bind to the molecule in a way that prevents the irreversible inhibition of the enzymes (Malbran et al, 1988). The exact mechanism by which these antibodies inactivate the C1-Inh is unknown and in order to better understand this disease, it is necessary to provide highly pure and fully functionally active C1-Inh in order to conduct further studies. However, of the many methods published for the purification of C1-Inh (Haupt et al, 1970; Harpel and Cooper, 1975; Reboul et al, 1977; Nilsson and Wiman, 1982; Harrison, 1983; Prograis et al, 1987), all are time-consuming and some do not yield highly pure protein, part of the reason being that the chromatography material was not very selective.
Recently, Hiemstra and collaborators (1987), have reported that jacalin binds C1-Inh in serum and causes complement activation. Jack fruit (Artocarous integrifolia) lectin, also called jacalin, is a galactosyl-specific lectin (Krishna Sastry et al, 1988) which can be used to separate human IgA.sub.1 from IgA.sub.2 (Gregory et al, 1987; Skea et al, 1988). Jacalin appears to bind only a small number of human serum proteins (Roque-Barreira and Campos-Neto, 1985).
Prior art purification procedures include several time-consuming chromatographic steps in which it is necessary to identify the C1-Inh containing fractions from each chromatographic step by either immunochemical or functional assays. Finally, dialysis of the C1-Inh containing pools is required between each step. All known procedures are tedious and time-consuming.